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Direct observation of strand passage by DNA-topoisomerase and its limited processivity.
- Source :
- PLoS ONE, Vol 7, Iss 4, p e34920 (2012)
- Publication Year :
- 2012
- Publisher :
- Public Library of Science (PLoS), 2012.
-
Abstract
- Type-II DNA topoisomerases resolve DNA entanglements such as supercoils, knots and catenanes by passing one segment of DNA duplex through a transient enzyme-bridged double-stranded break in another segment. The ATP-dependent passage reaction has previously been demonstrated at the single-molecule level, showing apparent processivity at saturating ATP. Here we directly observed the strand passage by human topoisomerase IIα, after winding a pair of fluorescently stained DNA molecules with optical tweezers for 30 turns into an X-shaped braid. On average 0.51 ± 0.33 µm (11 ± 6 turns) of a braid was unlinked in a burst of reactions taking 8 ± 4 s, the unlinked length being essentially independent of the enzyme concentration between 0.25-37 pM. The time elapsed before the start of processive unlinking decreased with the enzyme concentration, being ~100 s at 3.7 pM. These results are consistent with a scenario where the enzyme binds to one DNA for a period of ~10 s, waiting for multiple diffusional encounters with the other DNA to transport it across the break ~10 times, and then dissociates from the binding site without waiting for the exhaustion of transportable DNA segments.
Details
- Language :
- English
- ISSN :
- 19326203
- Volume :
- 7
- Issue :
- 4
- Database :
- Directory of Open Access Journals
- Journal :
- PLoS ONE
- Publication Type :
- Academic Journal
- Accession number :
- edsdoj.89ad52a83894c7898d797182ac8eb45
- Document Type :
- article
- Full Text :
- https://doi.org/10.1371/journal.pone.0034920